Large conductance, calcium and voltage-activated potassium (BK) channel is a unique member of potassium channel family, which has the largest single channel conductance and is dually activated by voltage and cytosolic free Ca2+ ([Ca2+]i), consisting of the pore-forming, voltage and Ca2+-sensing -subunits (BK) either alone or in association with tissue specific regulatory -subunits (1 - 4). BK channels are generally hard to open, requiring coincident membrane depolarization and [Ca2+]i rise for activation in vivo. However, with proteomic and electrophysiological approaches, we have recently identified a novel BK channel auxiliary subunit, leucine-rich repeat (LRR) containing membrane protein LRRC26 that causes an unprecedented large negative shift (~ -150 mV) in voltage dependence, allowing BK channel activation at even near resting voltages and calcium levels in excitable and non-excitable cells. LRRC26 represents a new family of BK channel auxiliary subunits, which is structurally distinct from the four known -subunits and designated as a -subunit. Like the presence of multiple tissue specific BK channel -subunits with different modulatory functions, we hypothesized that there exist other members of the LRRC26-like -subunits, which differentially modulate BK channels over a wide range of different cell types. The following two specific aims are designed to delineate the family members and to define the structural and functional features of the BK channel -subunits: 1) Identify and characterize new members of BK channel -subunits; 2) identify the structural determinants for modulatory function of the BK channel -subunits. We have identified five LRRC26 paralogs, LRRC38, LRRC52, LRRC55, LRTM1 and LRTM2, from protein database. Experiments are designed to determine the modulatory effects of these LRRC26 paralogs on BK channel function, and to identify key structural elements responsible for a LRR protein to be a BK channel modulator. Overall, the proposed research is designed to identify new family members and to define the structural and functional features of the BK channel -subunits. The findings from the proposed research may establish a broad LRR family of ion channel auxiliary subunits, gain an in-depth understanding of the LRRC26's unique capacity in BK channel modulation, and provide new protein targets for BK channel related disease treatment and drug development.